1. Field of the Invention
This invention relates to a scanning microscope for performing a confocal scanning operation.
2. Description of Related Art
For microscopes of this type, as discussed by, for example, T. R. Corle and G. S. Kino, "Confocal Scanning Optical Microscopy and Related Imaging Systems", Academic Press (1966), various microscopes have been proposed and manufactured. In view of types of confocal scanning devices used in such microscopes, the microscopes are roughly divided into two types, those in which a single-beam scanning device is used and those in which a multibeam scanning device is used. The fundamental constructions of these two types of microscopes will be explained below.
In the beginning, a description is given of the fundamental construction of a confocal microscope using the single-beam scanning device (which is hereinafter referred to as a single-beam scanning microscope) with reference to FIG. 1. A light beam emitted from a light source 1 is brought to a focus by means of a condenser lens 2, and an image of light transmitted through a first minute aperture 3 placed at this focus position is formed, through a first relay lens 4 and an objective lens 5, on a specimen 6. The light beam reflected by the specimen 6 is deflected in a direction different from that of the first minute aperture 3 by a beam splitter 7 interposed between the objective lens 5 and the first relay lens 4. This deflected beam is condensed, by a second relay lens 8, at a second minute aperture 9 located at a position conjugate with the first minute aperture 3 so that the intensity of light transmitted through this aperture is detected by a detector 10 such as a photomultiplier.
A scanning device 11 is disposed between the objective lens 5 and the beam splitter 7. The scanning device 11 has, for example, a galvanomirror or polygonal mirror to scan the image of the first minute aperture 3 on the surface of the specimen 6. A controller 12 connected to the scanning device 11 and the detector 10 is adapted to detect the position of image formation of the first minute aperture 3 on the specimen 6 in accordance with the amount of deflection of the light beam caused by the scanning device 11, and to obtain a specimen image in a wide region in accordance with a detection signal thereof and an output signal from the detector 10 relative to the position of image formation. In this way, the specimen image is displayed on an image output device 13 such as a TV monitor. Also, where a single-mode oscillation laser is used as the light source, the first minute aperture 3 is often removed.
Subsequently, as disclosed in U.S. Pat. No. 5,022,743, the fundamental construction of the confocal microscope in the case where a Nipkow disk is used as an example of the multibeam scanning device is explained with reference to FIG. 2. A light beam emitted from a light source 21 is radiated on a Nipkow disk 23 by a condenser lens 22. A plurality of beams which are split by passing through a plurality of small apertures provided in the Nipkow disk 23 are focused on the specimen 6 through a relay lens 24 and the objective lens 5. The light beams reflected from the specimen 6 are focused again on the small apertures of the Nipkow disk 23 through the objective lens 5 and the relay lens 24.
By a beam splitter 25 interposed between the Nipkow disk 23 and the condenser lens 22. light transmitted through the Nipkow disk 23 is deflected in a direction different from that of the light source 21, and is focused through a photographic lens 26 on an image sensor 27. Consequently, an image proportional to the reflectance of the specimen 6 is formed on the image sensor 27 and displayed on the image output device 13. The plurality of small apertures provided in the Nipkow disk 23 are spaced at prescribed intervals, and thus illumination light striking the specimen 6 at a time provides multiple spot illumination. However, since the Nipkow disk 23 is rotated at high speed by a motor 28, the entire surface of the specimen can be scanned in less time and a confocal observation with the naked eye is possible.
In this way, as seen from a difference in fundamental construction between both microscopes, the single-beam scanning microscope is high in illumination efficiency of the light source, but requires much time to scan the entire field of the microscope. Thus, it is impossible to make real-time observation at a video rate. In contrast to this, a confocal microscope using the multibeam scanning device (which is hereinafter referred to as a multibeam scanning microscope), not to speak of the microscope using the Nipkow disk, materially reduces the scanning time and thus is capable of making real-time observation at the video rate, which is very convenient. The present invention is directed to the confocal microscope using the multibeam scanning device, and prior art examples corresponding thereto are set forth in U.S. Pat. No. 5,022,743 mentioned above, U.S. Pat. No. 5,428,475, and Japanese Patent Preliminary Publication No. Hei 8-211296.
However, the multibeam scanning microscope which is suitable for such real-time observation has various problems. One of these problems refers to illumination efficiency. Specifically, when the Nipkow disk is used as the scanning device, as mentioned above, most of the light emitted from the light source is blocked by a portion, devoid of the small apertures, of the surface of the Nipkow disk, and hence illumination efficiency is so considerably impaired that fluorescence observation is not virtually made. Thus, some techniques for obviating this defect are proposed, and one of them is disclosed in U.S. Pat. No. 5,428,475 mentioned above.
According to this technique, a laser is used as a light source, and a light-collecting means such as a Fresnel lens array is provided between a condenser lens and a beam splitter. By doing so, light from the laser is split so that the light is collected at respective positions of the apertures of the Nipkow disk, and the light-collecting means is rotated together with the Nipkow disk by a motor. With this technique, the illumination efficiency is considerably improved and the fluorescence observation becomes possible, but there is the problem that it is very difficult to constitute the light-collecting means integrated with the Nipkow disk.
In the multibeam scanning microscope of the prior art, including the use of the Nipkow disk, locations on the specimen correspond individually to light-receiving elements. Thus, in order to obtain the specimen image while holding a resolving power governed by the optical system of the microscope, an image sensor with a high integration density of pixels, such as a CCD, has often been used as a light-receiving means. The image sensor, however, is such that as the integration density becomes high, a light-receiving area per unit pixel reduces, with a resulting decrease in sensitivity. Hence, the multibeam scanning microscope, in contrast with the single-beam scanning microscope, has the problem that detection with high sensitivity cannot be obtained. Furthermore, since the power consumption of the image sensor is proportional to the number of light-receiving elements, there is the problem that as the integration density of the image sensor becomes high, its power consumption increases. Consequently, it is considered that a high-sensitivity image pickup tube whose power consumption is small is used instead of the image sensor, but in this case, the problem arises that the image pickup tube is inferior in resolving power to the image sensor.
For multibeam scanning microscopes in which Nipkow disks are not used, many proposals are made. For example, Hei 8-211296 described above discloses a microscope designed so that a light source means including a matrix of a plurality of point sources and a detection means placed at a position conjugate with the point sources are used, and these point sources are periodically turned on and off to thereby scan a specimen and eliminate mechanical moving parts. However, even with this prior art example, since most of light emitted from the light source means is blocked and only a scanning spot part is used as illumination light, illumination efficiency is remarkably impaired. For the light-receiving means, like the above case, the image sensor with a high integration density of pixels must be used and a light-receiving sensitivity cannot be improved. Moreover, there is the problem that it is difficult to align a high integration density of point sources with the detection means.